Figure 7. CLASP2 regulates synaptic structure and synaptic protein levels. A, Ultrastructural analysis by electron microscopy of synaptic structure in hippocampal neuronal cultures infected with control and overexpressed human CLASP2 at 14 DIV. Representative images of asymmetric excitatory synapses of control and CLASP2 infected neurons. Scale bar, 500 nm. B, Hippocampal neuronal processes infected with control, human CLASP2, CLASP2 shRNA, or CLASP2 shRNA coexpressing human CLASP2 at 14 DIV stained with synapsin (Syn; green) and PSD95 (red). Scale bar, 4 μm. C, D, CLASP2 increases overall synapse number and area of colocalization of synapsin and PSD95 in neuronal processes at 14 DIV (*p < 0.05, ***p < 0.0001). E, Western blotting of CLASP2, various synaptic markers, and β-tubulin in response to control, human CLASP2, CLASP2 shRNA, or CLASP2 shRNA coexpressing human CLASP2 (rescue) in cortical neurons. F, We detected a significant increase of surface (S) compared with total (T) endogenous GluA1 receptor subunit in cultured cortical neurons that overexpressed CLASP2 at 14 DIV using a biotinylation assay (left, immunoblot; right, quantification). Total GluA1 levels were determined from input lysates, whereas surface GluA1 was determined after elution from NeutrAvidin precipitation (*p = 0.04).